Antigen presenting cells (APCs) play a central role in the initiation and expansion of cellular immune responses to cancer and virus infected cells. Foreign antigens are processed by APCs, which then interact with T lymphocytes or other effector cells. Part of the interaction pathway is via co-stimulatory molecules and ligands on APC and T cell surfaces. Currently, numerous clinical trials have been undertaken to test the feasibility and efficacy using antigen presenting cells (APCs) for therapeutic purposes. There are many types of APCs [1,2]. Different types of APCs differ in the origin, concentration in circulation, and the capacity to present antigens (Steinman, et al., Ann. Rev. Immunol. 9: 271 (1991); Caux et al., Immunol Today. 16(1):2-4 (1995); Hart and Mckenzie, Int Rev Immunol. 6(2-3):127-38 (1990); Austyn, Immunology 62(2):161-70 (1987)). The goal of vaccine therapy is to collect APCs or precursors, activate the cells using tumor antigen(s), and infuse the antigen-loaded cells to stimulate a T-cell immune in the patients that is sufficiently potent to cause regression of tumor. APCs have limited capability to kill tumor cells directly, but they can stimulate effector cells such as cytotoxic T cells and natural killer cells to destroy tumor cells. The most common procedure involves harvesting of immature APCs from blood, culturing ex vivo to activate the cells against tumor antigens, and infusing the activated cells to the same patient. Compositions and processes for ex vivo culture of APCs are disclosed in U.S. Pat. No. 6,121,044, incorporated herein by reference.
Blood APCs have several processing characteristics that make them ideal for development of commercial APC-based vaccines. They require only short-time culture (approximately 1 to 2 days) compared to the 7 days or more for APCs derived from monocytes or stem cells, and they do not require cytokines such as IL-4 or GM-CSF for maturation. Both of these characteristics make blood APCs substantially less complex and expensive to produce [3].
Two general processes occur during preparation of ex vivo antigen-loaded APCs for immunotherapy: (1) antigen processing; and (2) APC maturation. The first step in preparing a APC-based vaccine from blood APCs is to harvest immature blood APCs by methods known in the art such as by a standard mononuclear cell leukapheresis. Immature APCs effectively take up and process antigen, but are ineffective at presenting antigen to T cells because they lack cell surface costimulatory molecules and the capacity to secrete immunomodulatory cytokines that are essential to optimize T-cell activation. Thus, immature APCs must mature or become activated before they can effectively stimulate a T-cell-mediated immune response. Accordingly, immature APCs are cultured with an antigen under conditions to yield activated APCs. Various means for antigen delivery are disclosed in U.S. Pat. Nos. 5,976,546 and 6,210,662, incorporated herein by reference. After the activated APCs are washed to remove any unprocessed antigen, the activated APCs are subjected to a series of quality control tests including a determination of the potency of the APCs. The potency of dendritic cells is typically measured by their capacity to activate T cells and induce T cell proliferation. Activation can be monitored by upregulation of cell surface markers such as CD40, CD54, CD80and CD86 or by increased functional potency using such assays as allogenic mixed lymphocyte reactions (alloMLR).
Current methods of measuring the potency of the APCs utilize five to seven day bioassays which do not provide for “real-time” assessment of product potency. Accordingly, there is a need for techniques for the rapid identification of the potency of activated APCs that have the ability to elicit an immune response. This will, in turn, ensure that sufficient therapeutic quantities of active cells are infused into the patients, which is of importance in the design of therapeutic strategies in relation to disease such as cancer.